Preparation of and formulation comprising a MEK inhibitor

ABSTRACT

The present invention relates to processes for preparing 6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic acid (2-hydroxyethyoxy)-amide, processes for preparing crystallized 6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic acid (2-hydroxyethyoxy)-amide, and intermediates useful therefore. Also provided herein are pharmaceutical compositions comprising this crystallized compound.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.15/053,441, filed Feb. 25, 2016, which is a continuation of U.S.application Ser. No. 14/974,655, filed Dec. 18, 2015, which is adivisional application of U.S. application Ser. No. 14/057,498, filedOct. 18, 2013, which claims priority to U.S. Provisional Application No.61/716,169, filed Oct. 19, 2012, which is incorporated herein byreference in its entirety.

FIELD OF INVENTION

Provided herein are processes for preparing6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, processes for preparing crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, and intermediates useful therefore. Alsoprovided herein are pharmaceutical compositions comprising thiscrystallized compound.

BACKGROUND

Growth factor-mediated proliferative signals are transmitted from theextracellular environment to the nucleus through several pathways,including the RAS/RAF/MEK pathway. The RAS/RAF/MEK kinase signaltransduction pathway is activated through initial extracellular bindingand stimulation of tyrosine receptor kinases (RTKs) by their respectivecognate ligands. Upon autophosphorylation of specific tyrosine residuesin the cytosolic domain of RTKs, the Grb2-Sos complex translocates tothe plasma membrane, and converts the inactive RAS*GDP to activeRAS*GTP. The interaction between the Grb2 docking protein and theactivated kinases or the phosphorylated receptor associated proteins ismediated by the Src Homology (SH2) domain of the signaling protein thatrecognizes specific phosphotyrosine sequences. RAS undergoes aconformational change upon guanosine 5′-triphosphate (GTP) binding andcauses the recruitment of RAF-1 to the cytoplasmic membrane where it isphosphorylated by several kinases and simultaneous disphosphorylated atkey residues by protein phosphatase-2B. Activated RAF phosphorylates themitogen-activated protein kinase (MEK) on two serine residues in theactivation loop, which results in the activation of this protein kinase.MEK then phosphorylates and activates extracellular signal-regulatedkinase (ERK), allowing its translocation to the nucleus where itphosphorylates transcriptional factors permitting the expression of avariety of genes.

The RAS/RAF/MEK signal transduction pathway is deregulated, oftenthrough mutations that result in ectopic protein activation, in roughly⅓ of human cancers. This deregulation in turn results in a wide array ofcellular changes that are integral to the etiology and maintenance of acancerous phenotype including, but not limited to, the promotion ofproliferation and evasion of apoptosis (Dhillon et al., Oncogene, 2007,26: 3279-3290). Accordingly, the development of small moleculeinhibitors of key members of the RAS/RAF/MEK signal transduction pathwayhas been the subject of intense effort within the pharmaceuticalindustry and oncology community.

MEK is a major protein in the RAS/RAF/MEK pathway, which signals towardcell proliferation and survival, and frequently activated in tumors thathave mutations in the RAS or RAF oncogenes or in growth receptortyrosine kinases. MEK is a key player in the RAS/RAF/MEK pathway as itis downstream of RAS and RAF. Despite being only rarely mutated incancer (Murugan et al., Cell Cycle, 2009, 8: 2122-2124; Sasaki et al.,J. Thorac. Oncol., 2010, 5: 597-600), inhibitors of the MEK1 and MEK2proteins have also been targeted for small molecule inhibition owing totheir central position within the RAS/RAF/MEK signal transductionpathway signaling cascade (Fremin and Meloche, J. Hematol. Oncol., 2010,3:8). Recently a potent MEK inhibitor failed to demonstrate efficacy inclinical trials in patients with advanced non-small cell lung cancer(Haura et al., Clin. Cancer Res., 2010, 16: 2450-2457). The reason forfailure in this trial is not clear.

6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (hereinafter, “Compound A”) is abenzimidazole compound that is a known potent and selective inhibitor ofthe MEK1 and MEK2 proteins, and useful in the treatment ofhyperproliferative diseases, particularly cancer, in mammals. Forexample, in a recently published Phase I study of 28 patients sufferingfrom unresectable, locally advanced or metastatic biliary cancer and whohad received ≤1 prior systemic therapy, oral Compound A treatment (60 mgtwice daily) resulted in 1 complete regression, 1 partial regression and11 stable disease diagnoses after at least 6 weeks of treatment (Finn etal., J. Clin. Oncol. 30, 2012 (Supplement 4, 2012 GastrointestinalCancers Symposium, Abstract No. 220). Compound A has also beendemonstrated to be effective in the treatment of patients with eitherBRAFV600 or NRAS-mutant melanoma (Ascierto et al., J. Clin. Oncol. 30,2012 (Supplement, 2012 ASCO Annual Meeting, Abstract No. 8511).

The compound, as well as a process for its preparation, is disclosed inPCT Pub. No. WO 03/077914. The manufacturing process for preparingCompound A is described in Example 18 of this document. Themanufacturing process described therein are, although suitable, regardedas disadvantageous for commercial production.

Due to the high potency of this benzimidazole compound, in particular asMEK inhibitor, there is a need for improved manufacturing methods ofsuch compounds. In particular, there is a need to provide processes thatfulfill one or more of the following criteria: scalable, safer, simpler,higher yielding and more economical when compared to known processes.

There also remains a need for new solid forms for the treatment ofcancer.

The present invention is directed to an improved process for theproduction of Compound A that is suitable for small scale or large scalemanufacture, and useful intermediates thereof. The present invention isfurther directed to a process for the production of crystallizedCompound A as well as a new pharmaceutical composition suitable foradministration of this crystallized compound. It has been surprisinglydiscovered that crystallized Compound A prepared according to theinventive processes has an improved purity profile and an improvedphysical morphology which is advantageous in pharmaceutical drugdevelopment and manufacture.

SUMMARY OF THE INVENTION

Provided herein are processes for preparing6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, processes for preparing crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, and intermediates useful therefore. Alsoprovided herein are pharmaceutical compositions comprising thiscrystallized compound.

In one aspect, provided herein is a process for preparing a compound6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (herein referred to as “Compound A”):

wherein the process comprising the steps of:

-   -   a) reacting a compound of Formula (I):

-   -   -   with a suitable base to form an intermediate; and

    -   b) reacting said intermediate with a compound of Formula (II):

-   -   -   to provide a compound of Formula (III):

or a hydrate thereof,

wherein P¹ is a protecting group;

-   -   c) dissolving said compound of Formula (III) or a hydrate        thereof in a suitable solvent or solvent system; and    -   d) deprotecting said compound of Formula (III) or a hydrate        thereof with a suitable deprotecting reagent,        -   wherein P¹ in each occurrence may be the same or different,            and is a suitable protecting group, to provide Compound A.

In another aspect, provided herein is a process for preparing a compoundof Formula (III):

or a hydrate thereof, wherein the process includes the steps of:

a) reacting a compound of Formula (I):

-   -   with a suitable base to form an intermediate; and

b) reacting the intermediate with a compound of Formula (II):

wherein P¹ is a protecting group, such that the compound of Formula(III) or a hydrate thereof is formed.

In certain embodiments of both processes, steps a) and b) are carriedout as a “one-pot” synthesis, wherein the intermediate of step a) isreacted with the compound of Formula (II) without first being isolatedfrom the reaction mixture of step a). In one particular embodiment, stepa) comprises reacting the compound of Formula (I) with the suitable baseto form Intermediate 1 (structure shown below), and step b) comprisesreacting Intermediate 1 with the compound of Formula (II) to form thecompound of Formula (III), or a hydrate thereof. In one embodiment,Intermediate 1 is not isolated from the reaction mixture of step a)prior to step b). In another embodiment, Intermediate 1 is part of asolution comprising solvents selected from the group consisting of DMFand THF.

In contrast, in other embodiments, the intermediate formed in step a) isisolated from the reaction mixture before reacting with the compound ofFormula (II). In one embodiment, step a) comprises reacting the compoundof Formula (I) with the suitable base to form an intermediate of Formula(V) (structure shown below), and isolating the intermediate from thereaction mixture; and step b) comprises reacting the intermediate ofFormula (V) with the compound of Formula (II) to form the compound ofFormula (III), or a hydrate thereof. As such, in this embodiment, stepsa) and b) are not carried out as a “one-pot” procedure but as separateproduction steps wherein the intermediate of step a) is isolated beforereaction with the compound of Formula (II) in step b). In one particularembodiment, step a) comprises reacting the compound of Formula (I) withthe suitable base followed by reacting with an acid to form anintermediate of Formula (V), and isolating the intermediate from thereaction mixture; and step b) comprises reacting the intermediate ofFormula (V) with the compound of Formula (II) to form the compound ofFormula (III), or a hydrate thereof. In one embodiment of the process,step a) comprises crystallizing and collecting the intermediate from thereaction mixture. In one embodiment, the intermediate is crystallizedand collected by filtration. This additional isolation step can beadvantageous as it removes starting materials and process impuritiesprior to the coupling reaction with the compound of Formula (II). Incertain embodiments of the processes, isolating the intermediate of stepa) improves synthesis yields.

In another aspect, there is provided a process for preparing acrystallized form of Compound A:

the process including the steps of:

-   -   a) dissolving Compound A in a solution comprising (i.) a solvent        system comprising an ether and optionally an alcohol, and (ii.)        water to provide a solution;    -   b) adding a seed crystal suspension to the solution to provide a        suspension mixture;    -   d) adding water to the suspension mixture to provide a treated        mixture; and    -   e) cooling the treated mixture, to provide the crystallized        6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic        acid (2-hydroxyethyoxy)-amide.

In another aspect, there is provided a process for preparing acrystallized form of Compound A:

the process including the steps of:

-   -   a) dissolving Compound A in a solution comprising (i.) a solvent        system comprising an ether and optionally an alcohol, and (ii.)        water to provide a solution;    -   b) adding a seed crystal suspension to the solution to provide a        suspension mixture;    -   c) cooling the suspension mixture to provide a cooled suspension        mixture;    -   d) adding water to the cooled suspension mixture to provide a        treated mixture; and    -   e) cooling the treated mixture, to provide the crystallized        6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic        acid (2-hydroxyethyoxy)-amide.

In another aspect, there is provided a crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide prepared in accordance with the processdescribed herein.

In another aspect, there is provided a compound of formula (I):

Said compound of formula (I) is useful as an intermediate compound forthe synthesis of Compound A in accordance with the present invention.

In another aspect, provided herein is a compound of formula (V):

Said compound of Formula (V) is useful as an intermediate compound forthe synthesis of Compound A in accordance with the present invention.

In another aspect, there is provided a compound of formula (IV):

or a hydrate thereof. In a preferred embodiment, the compound of formula(IV) is in the form of its monohydrate. Said compound of formula (IV),including the monohydrate, is useful as an intermediate compound for thesynthesis of Compound A in accordance with the present invention.

In another aspect, there is provided a pharmaceutical compositioncomprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, at least one sugar, and at least onecellulose-derivative excipient.

In another aspect, there is provided a method of treating aproliferative disease, particularly cancer, in a subject, comprisingadministering to the subject in need thereof the pharmaceuticalcomposition comprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, at least one sugar, and at least onecellulose-derivative excipient.

DETAILED DESCRIPTION OF THE FIGURES

FIG. 1 shows two microscopy images of the agglomerated drug substanceCompound A produced by prior processes.

FIG. 2 shows a microscopy image of the crystalline Compound A producedby the new crystallization process of the present invention: (a) withoutmilling, (b) with jet-milling, and (c) with pin-milling.

DETAILED DESCRIPTION OF THE INVENTION

Provided herein are processes useful for the preparation and synthesisof a potent and selective MEK 1/2 inhibitor6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (“Compound A”), and useful intermediatesthereof. The present invention is further directed to a process for thesynthesis of crystallized Compound A as well as a pharmaceuticalcomposition suitable for administration of this crystallized compound.

These processes are advantageous over previously known processes (e.g,WO 03/077914) in several ways. For example, the instant processes forthe formation of Compound A have an improved purity profile with lowlevels (less than 1 ppm) of palladium.

The general terms used herein are defined with the following meanings,unless explicitly stated otherwise.

“Subject” for the purposes of the present invention includes humans andother animals, particularly mammals, and other organisms. Thus themethods are applicable to both human therapy and veterinaryapplications. In a preferred embodiment the patient is a mammal, and ina most preferred embodiment the patient is human.

The terms “effective amount” or “pharmaceutically effective amount” or“therapeutically effective amount” refer to a sufficient amount of anagent to provide the desired biological, therapeutic, and/orprophylactic result. That result can be reduction, amelioration,palliation, lessening, delaying, and/or alleviation of one or more ofthe signs, symptoms, or causes of a disease, or any other desiredalteration of a biological system. In reference to cancer, an effectiveamount comprises an amount sufficient to cause a tumor to shrink and/orto decrease the growth rate of the tumor (such as to suppress tumorgrowth) or to prevent or delay other unwanted cell proliferation. Insome embodiments, an effective amount is an amount sufficient to delaydevelopment. In some embodiments, an effective amount is an amountsufficient to prevent or delay recurrence. An effective amount can beadministered in one or more administrations. The effective amount of thedrug or composition may: (i) reduce the number of cancer cells; (ii)reduce tumor size; (iii) inhibit, retard, slow to some extent, andpreferably stop cancer cell infiltration into peripheral organs; (iv)inhibit (i.e., slow to some extent and preferably stop) tumormetastasis; (v) inhibit tumor growth; (vi) prevent or delay occurrenceand/or recurrence of tumor; and/or (vii) relieve to some extent one ormore of the symptoms associated with the cancer.

Unless otherwise indicated, “treating” or “treatment” of a disease,disorder, or syndrome, as used herein, means inhibiting the disease,disorder, or syndrome, that is, arresting its development; and relievingthe disease, disorder, or syndrome, that is, causing regression of thedisease, disorder, or syndrome. As is known in the art, in the contextof treatment, adjustments for systemic versus localized delivery, age,body weight, general health, sex, diet, time of administration, druginteraction and the severity of the condition may be necessary, and willbe ascertainable with routine experimentation by one of ordinary skillin the art.

“Prevention” means preventing the disease, disorder, or syndrome fromoccurring in a human, i.e. causing the clinical symptoms of the disease,disorder, or syndrome not to develop in an animal that may be exposed toor predisposed to the disease, disorder, or syndrome but does not yetexperience or display symptoms of the disease, disorder, or syndrome.

“Pharmaceutical composition” means a mixture or solution containing atleast one therapeutic agent to be administered to a subject, e.g., amammal or human, in order to prevent or treat a particular disease orcondition affecting the mammal.

“Pharmaceutically acceptable” means those compounds, materials,compositions and/or dosage forms, which are, within the scope of soundmedical judgment, suitable for contact with the tissues a subject, e.g.,a mammal or human, without excessive toxicity, irritation allergicresponse and other problem complications commensurate with a reasonablebenefit/risk ratio.

The terms “comprising” and “including” are used herein in theiropen-ended and non-limiting sense unless otherwise noted.

The terms “a” and “an” and “the” and similar references in the contextof describing the invention (especially in the context of the followingclaims) are to be construed to cover both the singular and the plural,unless otherwise indicated herein or clearly contradicted by context.Where the plural form is used for compounds, salts, and the like, thisis taken to mean also a single compound, salt, or the like.

As used herein, the terms “approximately” or “about” generally indicatea possible variation of no more than 10%, 5%, or 1% of a value.

As used herein, the term “isolated” is meant that a compound isseparated from the reaction mixture in which it is formed or detected.An isolated compound comprises less than 15%, less than 10%, less than6%, or less than 3% by weight of organic solvents or water. Non-limitedexamples of the separation methods include filtration, centrifugation,vacuum drying, precipitation, crystallization, and columnchromatography.

In one aspect, provided herein is a process for preparing a compound6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (herein referred to as “Compound A”)

wherein the process comprising the steps of:

-   -   a) reacting a compound of Formula (I):

-   -   -   with a suitable base to form an intermediate; and

    -   b) reacting said intermediate with a compound of Formula (II):

-   -   -   to provide a compound of Formula (III):

-   -   or a hydrate thereof,    -   wherein P¹ is a protecting group;    -   c) dissolving said compound of Formula (III) or a hydrate        thereof in a suitable solvent or solvent system; and    -   d) deprotecting said compound of Formula (III) or a hydrate        thereof with a suitable deprotecting reagent,        -   wherein P¹ in each occurrence may be the same or different,            and is a suitable protecting group, to provide Compound A.

In the process of step a), the compound of Formula (I) is reacted with asuitable base to produce an intermediate. Examples of suitable bases forthe foregoing reaction include, but are not limited to, sodiumhydroxide, potassium hydroxide, caesium hydroxide, lithium hydroxide,potassium trimethylsilanolate, lithium trimethylsilanolate, and sodiumtrimethylsilanolate. In a preferred embodiment, the suitable base ispotassium trimethylsilanolate. In another preferred embodiment, thesuitable base is sodium hydroxide.

The process of step a), in which a compound of Formula (I) is reactedwith a suitable base, may be performed in any suitable solvent orsolvent system. Suitable solvents include polar aprotic solvents such asacetone, acetonitrile, N,N-dimethylformamide, dimethylsulfoxide, ethylacetate, tetrahydrofuran, 2-methyl-tetrahydrofuran, and 1,4-dioxane.Suitable solvent systems include any combination of suitable solvents.In a preferred embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and THF. Suitable solvent systems can also includeone or more suitable solvents in combination with water. In oneparticular embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and water.

In one embodiment of the process described above, steps a) and b) arecarried out as a “one-pot” synthesis, wherein the intermediate of stepa) is reacted with the compound of Formula (II) without first beingisolated from the reaction mixture of step a) (see, e.g., Example 2A).In one embodiment, the intermediate of step a) is Intermediate 1:

In one particular embodiment of the process comprising the “one-pot”synthesis, step a) comprises reacting the compound of Formula (I) withthe suitable base to form Intermediate 1, and step b) comprises reactingIntermediate 1 with the compound of Formula (II) to form the compound ofFormula (III), or a hydrate thereof. In one embodiment, Intermediate 1is not isolated from the reaction mixture of step a) prior to step b).In another embodiment, Intermediate 1 is part of a solution comprisingsolvents selected from the group consisting of DMF and THF.

In another embodiment of the process, steps a) and b) are carried outwith isolation of the intermediate of step a) from the reaction mixtureprior to reacting with the compound of Formula (II). In one embodiment,the intermediate of step a) is a compound of Formula (V):

In another embodiment of the process wherein the intermediate of step a)is isolated from the reaction mixture of step a) prior to the reactionof step b), the process comprises isolating the intermediate (e.g., thecompound of Formula (V)) from the reaction mixture of step a) prior tostep b). In one embodiment, the isolation process comprisescrystallizing and collecting the intermediate (e.g., the compound ofFormula (V)) from the reaction mixture of step a) prior to step b). Inan embodiment, the intermediate is crystallized and collected byfiltration.

In another embodiment, step a) comprises reacting the compound ofFormula (I) with the suitable base to form an intermediate of Formula(V), and isolating the intermediate from the reaction mixture; and stepb) comprises reacting the intermediate of Formula (V) with the compoundof Formula (II) to form the compound of Formula (III), or a hydratethereof. In one particular embodiment of the process wherein theintermediate of step a) is isolated from the reaction mixture of step a)prior to the reaction of step b), step a) comprises reacting thecompound of Formula (I) with the suitable base followed by reacting withan acid to form the intermediate of Formula (V), and isolating theintermediate of Formula (V) from the reaction mixture; and step b)comprises reacting the intermediate of Formula (V) with the compound ofFormula (II) to form the compound of Formula (III), or a hydratethereof. In one embodiment, the acid is hydrochloric acid. In anotherembodiment, step a) of the process comprises crystallizing andcollecting the intermediate of Formula (V) from the reaction mixture ofstep a). In one embodiment, the intermediate is crystallized andcollected by filtration.

As used herein, the term “protecting group” is intended to refer tothose groups used to prevent reactive groups (such as carboxy, amino,hydroxy, and mercapto groups) from undergoing undesired reactions. Inparticular, suitable protecting groups for P¹ as used throughout theapplication include acid-labile protecting groups. Illustrative examplesof suitable acid-labile protecting groups for P¹ as used throughout theapplication include, but are not limited to: alkyl groups, such astertiary alkyls (e.g., tertiary C₄-C₇ alkyls such as t-butyl or tertiaryamyl); alkenyl groups; tertiary aryl-alkyl groups, such as1-methyl-1-phenylethyl (cumyl) or triphenylmethyl (trityl); groups thatresult in acetals, such as methoxymethyl, 1-ethoxyethyl,2-tetrahydropyranyl or 2-tetrahydrofuranyl; and silyl groups, such astrimethylsilyl, triethylsilyl or tert-butyl-dimethyldilyl. In apreferred embodiment, P¹ is t-butyl.

The process of step b), in which the intermediate from step a) isreacted with a compound of Formula (II), may be performed in thepresence of any coupling agent and a proton source.

Suitable proton sources include, but are not limited to, imidazolehydrochloride, pyridinium hydrochloride, triethylamine hydrochloride,N-methylmorpholine hydrochloride, and sulfonic acids such as e.g.,methanesulfonic acid, and preferably imidazole hydrochloride. Suitablecoupling agents include, but are not limited to1,1′-carbonyldiimidazole, isobutylchloroformate, pivaloyl chloride,oxalyl chloride, thionyl chloride, 1-propanephosphonic acid cyclicanhydride, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride (EDCI), and preferably 1,1′-carbonyldiimidazole. In thepreferred embodiment of the present invention, the process of step b) isperformed in the presence of the coupling agent 1,1′-carbonyldiimidazoleand the proton source imidazole hydrochloride. It is within theknowledge of one of ordinary skill in the art to optimize the process ofthe present invention for coupling agents other than1,1′-carbonyldiimidazole and proton sources other than imidazolehydrochloride.

The process of step c), in which the compound of Formula (III) or ahydrate thereof is dissolved in a suitable solvent or solvent system anddeprotected, may be performed in any suitable solvent or solvent system.Examples of suitable solvents include (a) polar protic solvents such asmethanol, ethanol, and isopropanol, and (b) polar aprotic solvents suchas acetone, acetonitrile, N,N-dimethylformamide, dimethylsulfoxide,ethyl acetate, and tetrahydrofuran. Suitable solvent systems include anycombination of suitable solvents.

In one embodiment, the reaction is performed in a polar aprotic solvent.In a preferred embodiment, the reaction is performed in acetonitrile.

The protecting group for P¹ may be removed using any suitabledeprotecting agent. Deprotection conditions for hydroxy protectinggroups will necessarily vary with the choice of protecting group. Alkylgroups or alkeynyl groups may be removed, for example, with an aqueousacid, such as phosphoric acid. Tertiary aryl-alkyl groups may beremoved, for example, by aqueous acid. Silyl groups may be removed, forexample, by fluoride or by aqueous acid. It would be understood by oneof ordinary skill that acid-labile protecting groups may be removed withan aqueous acid. Suitable deprotection agents for those protectinggroups illustrated above may include, but are not limited to, an aqueousacid such as phosphoric acid, hydrochloric acid or sulfuric acid;non-aqueous acids such as hydrogen chloride acid in isopropyl alcohol orother suitable organic solvents such as 1,4-dioxane or tetrahydrfuran,trimethylsilylchloride, trifluoroacetic acid or p-Toluenesulfonic acid.

In one embodiment, when protecting group P¹ is t-butyl, a suitabledeprotection agent may be selected from an aqueous acid such asphosphoric acid, hydrochloric acid or sulfuric acid; 5 M hydrochloricacid in isopropyl alcohol, trimethylsilylchloride, or p-Toluenesulfonicacid (hydrate). Preferably, when protecting group P¹ is t-butyl, thesuitable deprotection agent is aqueous phosphoric acid.

In one embodiment, when the protecting group P¹ is t-butyl, the suitablesolvent or solvent system in step c) is selected from acetonitrile,tetrahydrofuran, methanol and ethanol, and the deprotection agent isphosphoric acid.

In a further embodiment, when the protecting group P¹ is t-butyl, thesuitable solvent or solvent system in step c) is selected fromacetonitrile, tetrahydrofuran, methanol, and ethanol, and thedeprotection agent is hydrochloric acid.

In a further embodiment of this aspect of the invention, step d) isfollowed by Compound A provided in step d) may be further converted intoany pharmaceutically acceptable salt thereof. A “pharmaceuticallyacceptable salt”, as used herein, unless otherwise indicated, includessalts of acidic and basic groups which may be present in the compoundsof the present invention. The compounds of the present invention thatare basic in nature are capable of forming a wide variety of salts withvarious inorganic and organic acids. The acids that may be used toprepare pharmaceutically acceptable acid addition salts of such basiccompounds of the present invention are those that form non-toxic acidaddition salts, i.e., salts containing pharmaceutically acceptableanions, such as the acetate, benzoate, bromide, chloride, citrate,fumarate, hydrobromide, hydrochloride, iodide, lactate, maleate,mandelate, nitrate, oxalate, salicylate, succinate, and tartrate salts.Since a single compound of the present invention may include more thanone acidic or basic moieties, the compounds of the present invention mayinclude mono, di or tri-salts in a single compound.

In the case of an acidic moiety in a compound of the present invention,a salt may be formed by treatment of a compound of the present inventionwith a basic compound, particularly an inorganic base. Preferredinorganic salts are those formed with alkali and alkaline earth metalssuch as lithium, sodium, potassium, barium and calcium. Preferredorganic base salts include, for example, ammonium, dibenzylammonium,benzylammonium, 2-hydroxyethylammonium, bis(2-hydroxyethyl)ammonium,phenylethylbenzylamine, dibenzyl-ethylenediamine, and the like salts.Other salts of acidic moieties may include, for example, those saltsformed with procaine, quinine and N-methylglusoamine, plus salts formedwith basic amino acids such as glycine, ornithine, histidine,phenylglycine, lysine and arginine. An especially preferred salt is asodium or potassium salt of a compound of the present invention.

With respect to basic moieties, a salt is formed by the treatment of acompound of the present invention with an acidic compound, particularlyan inorganic acid. Preferred inorganic salts of this type may include,for example, the hydrochloric, hydrobromic, sulfuric, phosphoric or thelike salts. Preferred organic salts of this type, may include, forexample, salts formed with cetic, succinic, citric, maleic, fumaric,D-glutamic, glycolic, benzoic, cinnamic and the like organic acids. Anespecially preferred salt of this type is a hydrochloride or sulfatesalt of Compound A.

It is understood that, in accordance with the present invention, a basemay be added to the acidic reaction mixture formed in deprotection stepd) to reach a pH in the range of 5-9. Preferably, a base is added toneutralize the acidic reaction mixture formed in deprotection step d) topH=approximately 8-8.5. Examples of suitable bases include, but are notlimited to, potassium hydroxide, sodium hydroxide, lithium hydroxide andammonium hydroxide. Preferably, the additional base is potassiumhydroxide.

In a preferred embodiment, the suitable base in step a) is potassiumtrimethylsilanolate, the protecting group P¹ is t-butyl, the suitablesolvent or solvent system of step c) is acetonitrile, and the suitabledeprotection agent in step d) is aqueous phosphoric acid.

In another preferred embodiment, the suitable base in step a) is sodiumhydroxide, the protecting group P¹ is t-butyl, the suitable solvent orsolvent system of step c) is acetonitrile, and the suitable deprotectionagent in step d) is aqueous phosphoric acid.

In another preferred embodiment, the suitable base in step a) ispotassium trimethylsilanolate, the protecting group P¹ is t-butyl, thesuitable solvent or solvent system of step c) is acetonitrile, thesuitable deprotection agent in step d) is phosphoric acid, and a furtherbase is added, wherein the further base is potassium hydroxide.

In still another preferred embodiment, the suitable base in step a) issodium hydroxide, the protecting group P¹ is t-butyl, the suitablesolvent or solvent system of step c) is acetonitrile, the suitabledeprotection agent in step d) is phosphoric acid, and a further base isadded, wherein the further base is potassium hydroxide.

In another aspect, provided herein is a process for preparing a compoundof Formula (III),

or a hydrate thereof, wherein the process includes the steps of:

a) reacting a compound of Formula (I):

-   -   with a suitable base to form an intermediate; and

b) reacting said intermediate with a compound of Formula (II):

wherein P¹ is a protecting group, such that the compound of Formula(III) or a hydrate thereof is formed.

The process of step a), in which the compound of Formula (I) is reactedwith a suitable base to produce an intermediate. Examples of suitablebases for the foregoing reaction include, but are not limited to, sodiumhydroxide, potassium hydroxide, caesium hydroxide, lithium hydroxide,potassium trimethylsilanolate, lithium trimethylsilanolate, and sodiumtrimethylsilanolate. In a preferred embodiment, the suitable base ispotassium trimethylsilanolate. In another preferred embodiment, thesuitable base is sodium hydroxide.

The process of step a), in which a compound of Formula (I) is reactedwith a suitable base, may be performed in any suitable solvent orsolvent system. Suitable solvents include polar aprotic solvents such asacetone, acetonitrile, N,N-dimethylformamide, dimethylsulfoxide, ethylacetate, tetrahydrofuran, 2-methyl-tetrahydrofuran, and 1,4-dioxane.Suitable solvent systems include any combination of suitable solvents.In a preferred embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and THF. Suitable solvent systems can also includeone or more suitable solvents in combination with water. In oneparticular embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and water.

In one embodiment of the process, steps a) and b) are carried out as a“one-pot” synthesis as described above. In one embodiment, theintermediate of step a) is Intermediate 1. In another embodiment, stepa) comprises reacting the compound of Formula (I) with the suitable baseto form Intermediate 1, and step b) comprises reacting Intermediate 1with the compound of Formula (II) to form the compound of Formula (III),or a hydrate thereof. In one embodiment, Intermediate 1 is not isolatedfrom the reaction mixture of step a) prior to step b). In anotherembodiment, Intermediate 1 is part of a solution comprising solventsselected from the group consisting of DMF and THF.

In another embodiment of the process, steps a) and b) are carried outwith isolation of the intermediate of step a) prior to the reaction ofstep b) as described above. In one embodiment, the intermediate of stepa) is a compound of Formula (V). In another embodiment, the processcomprises isolating the intermediate (e.g., the compound of Formula (V))from the reaction mixture of step a) prior to step b). In oneembodiment, the isolation process comprises crystallizing and collectingthe intermediate (e.g., the compound of Formula (V)) from the reactionmixture of step a) prior to step b). In one embodiment, the intermediateis crystallized and collected by filtration.

In another embodiment of the process wherein the intermediate of step a)is isolated from the reaction mixture of step a) prior to the reactionof step b), step a) comprises reacting the compound of Formula (I) withthe suitable base to form an intermediate of Formula (V), and isolatingthe intermediate from the reaction mixture; and step b) comprisesreacting the intermediate of Formula (V) with the compound of Formula(II) to form the compound of Formula (III), or a hydrate thereof. In oneparticular embodiment, step a) comprises reacting the compound ofFormula (I) with the suitable base followed by reacting with an acid toform the intermediate of Formula (V), and isolating the intermediate ofFormula (V) from the reaction mixture; and step b) comprises reactingthe intermediate of Formula (V) with the compound of Formula (II) toform the compound of Formula (III), or a hydrate thereof. In oneembodiment, the acid is hydrochloric acid.

Examples of suitable protecting groups P¹ include those disclosed above,wherein the text is hereby incorporated by reference in its entirety.

The process of step b), in which the intermediate from step a) isreacted with a compound of Formula (II), may be performed in thepresence of any coupling agent and a proton source. Suitable protonsources include, but are not limited to, imidazole hydrochloride,pyridinium hydrochloride, triethylamine hydrochloride,N-methylmorpholine hydrochloride, and sulfonic acids such as e.g.,methanesulfonic acid, and preferably imidazole hydrochloride. Suitablecoupling agents include, but are not limited to1,1′-carbonyldiimidazole, isobutylchloroformate, pivaloyl chloride,oxalyl chloride, thionyl chloride, 1-propanephosphonic acid cyclicanhydride, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride (EDCI), and preferably 1-1′-carbonyldiimidazole. In thepreferred embodiment of the present invention, the process of step b) isperformed in the presence of the coupling agent 1,1′-carbonyldiimidazoleand the proton source imidazole hydrochloride. It is within theknowledge of one of ordinary skill in the art to optimize the process ofthe present invention for coupling agents other than1,1′-carbonyldiimidazole and proton sources other than imidazolehydrochloride.

In another aspect of the invention, a new process for preparing a newcrystallized form of Compound A has been discovered. Prior synthesisprocesses of Compound A or pharmaceutically acceptable salts thereof,e.g., those in WO03/077914, have been demonstrated to possess thefollowing key disadvantages for pharmaceutical drug development: (a) thesynthesized drug substance typically formed big lumps (agglomerates) ofpowder, (b) insufficient purity profile and yield, and (c) thesynthesized drug substance had a “sticky” morphology with poorflowability. Prior processes would produce highly agglomerated materialof Compound A which would build lumps, with some having a diameter up to15 mm. These issues are significant issues that hinder the reliable,repeatable, and controlled large-scale production of pharmaceuticalcompositions comprising Compound A or pharmaceutically acceptable saltsthereof.

A new process for preparing a crystallized form of Compound A has beendiscovered that surprisingly produces a crystalline form of Compound Ahaving significantly improved purity profile and improved physicalmorphology (e.g., reduction of sticky crystals/particles, improvedflowability). Compound A has been found to have very low solubility inmost standard solvents (i.e., less than 1% at room temperature). Due tothis low solubility, it is difficult to perform crystallization bystandard cooling methods and to control crystal growth. However, it hasbeen discovered that water, which is in general an anti-solvent(solubility <0.01% at a broad temperature range, also unexpectedly actsas a solvent for Compound A when used in a novel solvent mixturecomprising an ether and optionally an alcohol, and thus significantlyincreases the solubility for Compound A. While a small additional ofanti-solvent to the solvent can typically slightly increase solubility,the addition of water to a solvent mixture of tetrahydrofuran andmethanol increased the solubility of Compound A by approximately 50%compared to the solvent mixture of methanol and tetrahydrofuran withoutwater. This improvement in solubility contributes to an improvement inthe purity profile of the final drug substance.

In addition, the new process for preparing a crystallized form ofCompound A produces a new crystalline form of Compound A with reducedsticky behavior and improved flowability. This improvement isdemonstrated with or without a subsequent milling step as discussedbelow. (See, FIG. 1 as compared to FIG. 2). The additional milling step,however, provides an advantageous further improvement to the stickybehavior and flowability of Compound A.

Thus, in one aspect, there is provided a process for preparing acrystallized form of Compound A:

the process including the steps of:

a) dissolving Compound A in a solution comprising (i.) a solvent systemcomprising an ether and optionally an alcohol, and (ii.) water toprovide a solution;

b) adding a seed crystal suspension to the solution to provide asuspension mixture;

d) adding water to the suspension mixture to provide a treated mixture;and

e) cooling the treated mixture, to provide the crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide.

In another aspect, there is provided a process for preparing acrystallized form of Compound A:

the process including the steps of:

-   -   a) dissolving Compound A in a solution comprising (i.) a solvent        system comprising an ether and optionally an alcohol, and (ii.)        water to provide a solution;    -   b) adding a seed crystal suspension to the solution to provide a        suspension mixture;    -   c) cooling the suspension mixture to provide a cooled suspension        mixture;    -   d) adding water to the cooled suspension mixture to provide a        treated mixture; and    -   e) cooling the treated mixture, to provide the crystallized        6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic        acid (2-hydroxyethyoxy)-amide.

In accordance with step a) of this aspect of the present invention,Compound A is dissolved in a solution comprising (i.) a solvent systemcomprising an ether and optionally an alcohol, and (ii.) water. It isunderstood that step a) may be performed by either: (a) adding CompoundA to a solution comprising (i.) a solvent system comprising an ether andoptionally an alcohol, and (ii.) water, or (b) adding each component ofthe solution to Compound A.

Suitable ethers include THF.

In accordance with the present invention, the solution of step a) doesnot require inclusion of an alcohol. A solution comprising both an etherand an alcohol, however, is preferred. Suitable alcohols include, butare not limited to methanol, ethanol, and isopropanol. Preferably, thealcohol is methanol.

In one embodiment, Compound A is dissolved in a solution consisting ofthe solvent THF and water.

In one embodiment, Compound A is dissolved in a solution comprising orconsisting of (i.) a solvent system including both an ether and analcohol, and (ii.) water. Preferred, the solution comprises a solventsystem including includes an alcohol and THF, and water.

In a preferred embodiment, Compound A is dissolved in a solutionconsisting of (i.) a solvent system consisting of methanol and THF, and(ii.) water.

The dissolution of Compound A in the solution of step a) is facilitatedby heating the mixture of Compound A and the solution comprising (i.) asolvent system comprising an ether and an alcohol, and (ii.) water to aninternal temperature of about 52-56° C. prior to addition of the seedcrystal suspension.

In one aspect, the mixture of Compound A and the solution comprising(i.) a solvent system comprising an ether and an alcohol, and (ii.)water is heated to an internal temperature of about 53-55° C.

The seed crystal suspension is added to the solution in step b) toprovide a suspension mixture. The solution is advantageously cooled (i.)after heating the mixture of Compound A and the solution comprising (i.)a solvent system comprising an ether and an alcohol, and (ii.) water toan internal temperature of about 52-56° C. and (ii.) before addition ofthe seed crystal suspension.

In one aspect, water is added to the suspension mixture to provide atreated mixture. In another aspect, the suspension mixture is cooled toa temperature of about 30-50° C. before water is added. In a preferredembodiment, the suspension mixture is cooled to a temperature of about47-48° C. before water is added.

As discussed above, the crystallization process makes use ofanti-solvent cooling system. In step d), water is added to thesuspension or the cooled suspension mixture to provide a treatedmixture. Water, which in general acts as an anti-solvent (solubility<0.01% over a wide temperature range), also unexpectedly acts as asolvent for Compound A when part of the water/methanol/tetrahydrofuransolvent system. Thus, adding water unexpectedly has the effect ofsignificantly increasing the solubility of Compound A. Using thisprocess, it is found that at 65° C., the maximum solubility of CompoundA is reached with a mixture of water (24%), methanol (38%), andtetrahydrofuran (38%), with solubility of Compound A decreasing withfurther addition of water.

The water can be added over a period of 5 to 35 hours, such that thewater does not exceed 70% w/w, preferably 65% (w/w), of the solventsystem. In one embodiment, the water is added over a period of 10-25hours, preferably 25 hours. In another embodiment, the water is addedover a period of 25 hours such that 33% is added within 15 hours, and66% is added within 10 hours.

In another aspect, the components in the solvent system upon completionof the addition of water to the suspension or the cooled suspensionmixture have a final ratio of alcohol/ether/water in the range between40/40/20 and 15/15/70 w/w, wherein w/w is referring to the weightpercentage of each component relative to the other components of thesolvent/antisolvent system. In a preferred embodiment, the final ratioof alcohol/ether/water is approximately 20/20/60 w/w (20/20/60 w/w).

In step e), crystallized Compound A is finally obtained by cooling thetreated mixture. The treated mixture is advantageously cooled over aperiod of 5 to 25 hours. In one aspect, the treated mixture is cooledover a period of 8 to 15 hours, 8 to 12 hours or 9 to 11 hours. In apreferred embodiment, the treated mixture is cooled over a period ofapproximately 10 hours. The treated mixture is advantageously cooled toan internal temperature of about 1-10° C., preferably about 3-5° C. In apreferred embodiment, the treated mixture is cooled to an internaltemperature of about 3-5° C. over a period of approximately 9-11 hours.

After crystallized Compound A is filtered, it can be dried, for example,under vacuum, or in a vacuum oven.

In a further embodiment, the crystallized Compound A provided in step e)in the inventive process above may be subsequently milled. Suitablemilling techniques would be known to one of ordinary skill and wouldinclude, but not be limited to, pin-milling or j et-milling.

Smaller primary particles often lead to higher agglomeration. However,after milling, crystallized Compound A unexpectedly showed a reducedstickiness, even after storage times of more than three months.

In one further aspect, there is provided is a process for preparing acrystallized form of Compound A comprising the steps of:

-   -   a) reacting a compound of Formula (I):

-   -   -   with a suitable base to form an intermediate; and

    -   b) reacting said intermediate with a compound of Formula (II):

-   -   -   to provide a compound of Formula (III):

-   -   -   or a hydrate thereof, wherein P¹ is a protecting group;

    -   c) dissolving said compound of Formula (III) or a hydrate        thereof in a suitable solvent or solvent system;

    -   d) deprotecting said compound of Formula (III) or a hydrate        thereof with a suitable deprotecting reagent,        -   wherein P¹ in each occurrence may be the same or different,            and is a suitable protecting group, to provide Compound A;

    -   e) dissolving Compound A in a solution comprising (i.) a solvent        system comprising an ether and optionally an alcohol, and (ii.)        water to provide a solution;

    -   f) adding a seed crystal suspension to the solution to provide a        suspension mixture;

    -   g) adding water to the suspension mixture to provide a treated        mixture; and

    -   h) cooling the treated mixture, to provide the crystallized        6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic        acid (2-hydroxyethyoxy)-amide.

In another aspect, there is provided is a process for preparing acrystallized form of Compound A comprising the steps of:

-   -   a) reacting a compound of Formula (I):

-   -   -   with a suitable base to form an intermediate; and

    -   b) reacting said intermediate with a compound of Formula (II):

-   -   -   to provide a compound of Formula (III):

-   -   -   or a hydrate thereof, wherein P¹ is a protecting group;

    -   c) dissolving said compound of Formula (III) or a hydrate        thereof in a suitable solvent or solvent system;

    -   d) deprotecting said compound of Formula (III) or a hydrate        thereof with a suitable deprotecting reagent, wherein P¹ in each        occurrence may be the same or different, and is a suitable        protecting group, to provide Compound A,

    -   e) dissolving Compound A in a solution comprising (i.) a solvent        system comprising an ether and optionally an alcohol, and (ii.)        water to provide a solution;

    -   f) adding a seed crystal suspension to the solution to provide a        suspension mixture; cooling the suspension mixture to provide a        cooled suspension mixture;

    -   g) adding water to the cooled suspension mixture to provide a        treated mixture; and

    -   h) cooling the treated mixture, to provide the crystallized        6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylic        acid (2-hydroxyethyoxy)-amide.

The process of step a), in which the compound of Formula (I) is reactedwith a suitable base to produce an intermediate. Examples of suitablebases for the foregoing reaction include, but are not limited to, sodiumhydroxide, potassium hydroxide, caesium hydroxide, lithium hydroxide,potassium trimethylsilanolate, lithium trimethylsilanolate, and sodiumtrimethylsilanolate. In a preferred embodiment, the suitable base ispotassium trimethylsilanolate. In another preferred embodiment, thesuitable base is sodium hydroxide.

The process of step a), in which a compound of Formula (I) is reactedwith a suitable base, may be performed in any suitable solvent orsolvent system. Suitable solvents include polar aprotic solvents such asacetone, acetonitrile, N,N-dimethylformamide, dimethylsulfoxide, ethylacetate, tetrahydrofuran, 2-methyl-tetrahydrofuran, and 1,4-dioxane.Suitable solvent systems include any combination of suitable solvents.In a preferred embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and THF. Suitable solvent systems can also includeone or more suitable solvents in combination with water. In oneparticular embodiment, the reaction is performed in a mixture ofN,N-dimethylformamide and water.

In one embodiment of the process, steps a) and b) are carried out as a“one-pot” synthesis as described above. In one embodiment, theintermediate of step a) is intermediate 1. In another embodiment, stepa) comprises reacting the compound of Formula (I) with the suitable baseto form Intermediate 1, and step b) comprises reacting Intermediate 1with the compound of Formula (II) to form the compound of Formula (III),or a hydrate thereof. In one embodiment, Intermediate 1 is a solutioncomprising solvents selected from the group consisting of DMF and THF.

In another embodiment of the process, steps a) and b) are carried outwith isolation of the intermediate of step a) prior to the reaction ofstep b) as described above. In one embodiment, the intermediate of stepa) is a compound of Formula (V). In another embodiment, the processcomprises isolating the intermediate (e.g., the compound of Formula (V))from the reaction mixture of step a) prior to step b). In oneembodiment, the process comprises crystallizing and collecting theintermediate (e.g., the compound of Formula (V)) from the reactionmixture of step a) prior to step b). In one embodiment, the intermediateis crystallized and collected by filtration.

In another embodiment of the process wherein the intermediate of step a)is isolated from the reaction mixture of step a) prior to the reactionof step b), step a) comprises reacting the compound of Formula (I) withthe suitable base to form an intermediate of Formula (V), and isolatingthe intermediate from the reaction mixture; and step b) comprisesreacting the intermediate of Formula (V) with the compound of Formula(II) to form the compound of Formula (III), or a hydrate thereof. In oneparticular embodiment, step a) comprises reacting the compound ofFormula (I) with the suitable base followed by reacting with an acid toform the intermediate of Formula (V), and isolating the intermediate ofFormula (V) from the reaction mixture; and step b) comprises reactingthe intermediate of Formula (V) with the compound of Formula (II) toform the compound of Formula (III), or a hydrate thereof. In oneembodiment, the acid is hydrochloric acid.

Suitable protecting groups and deprotecting reagents of steps b) and d)are provided above, and hereby incorporated by reference in itsentirety.

The process of step b), in which the intermediate from step a) isreacted with a compound of Formula (II), may be performed in thepresence of any coupling agent and a proton source. Suitable protonsources include, but are not limited to, imidazole hydrochloride,pyridinium hydrochloride, triethylamine hydrochloride,N-methylmorpholine hydrochloride, and sulfonic acids such as e.g.,methanesulfonic acid, and preferably imidazole hydrochloride. Suitablecoupling agents include, but are not limited to,1,1′-carbonyldiimidazole, isobutylchloroformate, pivaloyl chloride,oxalyl chloride, thionyl chloride, 1-propanephosphonic acid cyclicanhydride, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride (EDCI), and preferably 1,1′-carbonyldiimidazole. In thepreferred embodiment of the present invention, the process of step b) isperformed in the presence of the coupling agent 1,1′-carbonyldiimidazoleand the proton source imidazole hydrochloride. It is within theknowledge of one of ordinary skill in the art to optimize the process ofthe present invention for coupling agents other than1,1′-carbonyldiimidazole and proton sources other than imidazolehydrochloride.

It is understood that, in accordance with the present invention, a basemay be added to the acidic reaction mixture formed in deprotection stepd) to reach a pH in the range of 5-9. Preferably, a base is added toneutralize the acidic reaction mixture formed in deprotection step d) topH=approximately 8-8.5. Examples of suitable bases include, but are notlimited to, potassium hydroxide, sodium hydroxide, lithium hydroxide andammonium hydroxide. Preferably, the additional base is potassiumhydroxide.

In accordance with step e) of this aspect of the present invention,Compound A is dissolved in a solution comprising (i.) a solvent systemcomprising an ether and optionally an alcohol, and (ii.) water. It isunderstood that step e) may be performed by either: (a) adding CompoundA to a pre-mixed solution comprising (i.) a solvent system comprising anether and optionally an alcohol, and (ii.) water, or (b) adding eachcomponent of the solution to Compound A.

Suitable ethers include THF.

In accordance with the present invention, the solution of step e) doesnot require inclusion of an alcohol. A solution comprising both an etherand an alcohol, however, is preferred. Suitable alcohols include, butare not limited to methanol, ethanol, and isopropanol. Preferably, thealcohol is methanol.

In a preferred embodiment, Compound A is dissolved in a solutionconsisting of (i.) a solvent system consisting of methanol and THF, and(ii.) water.

The dissolution of Compound A in the solution of step e) is facilitatedby heating the mixture of Compound A and the solution comprising (i.) asolvent system comprising an ether and an alcohol, and (ii.) water to aninternal temperature of about 52-56° C. prior to addition of the seedcrystal suspension. In one aspect, the mixture of Compound A and thesolution comprising (i.) a solvent system comprising an ether and analcohol, and (ii.) water is heated to an internal temperature of about53-55° C.

The seed crystal suspension is added to the solution in step f) toprovide a suspension mixture. The solution is advantageously cooled (i.)after heating the mixture of Compound A and the solution comprising (i.)a solvent system comprising an ether and an alcohol, and (ii.) water toan internal temperature of about 52-56° C. and (ii.) before addition ofthe seed crystal suspension.

In one aspect, water is added to the suspension mixture to provide atreated mixture. In another aspect, the suspension mixture is cooled toa temperature of about 30-50° C. before water is added. In a preferredembodiment, the suspension mixture is cooled to a temperature of about47-48° C. before water is added.

In step g), the water can be added over a period of 5 to 35 hours, suchthat the water does not exceed 70% w/w, preferably 65% (w/w), of thesolvent system. In one embodiment, the water is added over a period of10-25 hours, preferably 25 hours. In another embodiment, the water isadded over a period of 25 hours such that 33% is added within 15 hours,and 66% is added within 10 hours.

In another aspect, the components in the solvent system upon completionof the addition of water to the suspension or the cooled suspensionmixture have a final ratio of alcohol/ether/water in the range between40/40/20 and 15/15/70 w/w, wherein w/w is referring to the weightpercentage of each component relative to the other components of thesolvent/antisolvent system. In a preferred embodiment, the final ratioof alcohol/ether/water is approximately 20/20/60 w/w (20/20/60 w/w).

In step h), crystallized Compound A is finally obtained by cooling thetreated mixture. The treated mixture is advantageously cooled over aperiod of 5 to 25 hours. In one aspect, the treated mixture is cooledover a period of 8 to 15 hours, 8 to 12 hours or 9 to 11 hours. In apreferred embodiment, the treated mixture is cooled over a period ofapproximately 10 hours.

The treated mixture is advantageously cooled to an internal temperatureof about 1-10° C., preferably about 3-5° C. In a preferred embodiment,the treated mixture is cooled to an internal temperature of about 3-5°C. over a period of approximately 9-11 hours. After crystallizedCompound A is filtered, it can be dried, for example, under vacuum, orin a vacuum oven.

Crystallized Compound A produced by the inventive process can be furthermilled (e.g., jet milling or pin milling).

In a further aspect, there is provided a crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide prepared in accordance with the processdescribed herein above, which is hereby incorporated by reference in itsentirety. It is understood that the crystallized Compound A includes thedrug substance prepared with or without further milling.

In another aspect, there is provided a compound of formula (I):

Said compound of formula (I) is useful as an intermediate compound forthe synthesis of Compound A in accordance with the present invention.

In another aspect, provided herein is a compound of Formula (V):

Said compound of Formula (V) is useful as an intermediate compound forthe synthesis of Compound A in accordance with the present invention.

In another aspect, there is provided a compound of formula (IV):

or a hydrate thereof. In a preferred embodiment, the compound of formula(IV) is in the form of its monohydrate. Said compound of formula (IV),including the monohydrate thereof, is useful as an intermediate compoundfor the synthesis of Compound A in accordance with the presentinvention.Pharmaceutical Compositions Comprising Crystallized Compound A

In another aspect, there is provided a pharmaceutical compositioncomprising crystallized Compound A and at least one pharmaceuticallyacceptable carrier or excipient. The pharmaceutical compositioncomprises crystallized Compound A, at least one sugar, and at least onecellulose-derivative excipient. The composition is particularly usefulfor the treatment of cancer in a subject in need thereof, preferablyhumans.

In the pharmaceutical compositions of the present invention, thecrystallized Compound A is in a crystal form produced by thecrystallization process described above, with or without the millingstep. In one embodiment of the pharmaceutical composition providedherein, the pharmaceutical composition comprises about 5-35%crystallized Compound A by weight of composition. In a furtherembodiment, the pharmaceutical composition comprises about 5-11%crystallized Compound A by weight of composition. In another preferredembodiment, the pharmaceutical composition comprises about 6.25%crystallized Compound A by weight of composition. In another preferredembodiment, the pharmaceutical composition comprises about 10%crystallized Compound A by weight of composition.

In another embodiment, the pharmaceutical composition comprisesapproximately 15 mg or 45 mg crystallized Compound A.

Suitable sugars for use in the pharmaceutical compositions include, butare not limited to, lactose (e.g., spray-dried lactose, lactosemonohydrate), maltose, fructose, galactose, confectioner's sugar,compressible sugar, dextrates, dextrin, dextrose, mannitol, Nu-Tab,Di-Pac, Emdex, and sucrose. In a preferred embodiment, the sugar used inthe pharmaceutical composition is lactose, particularly lactosemonohydrate.

In one embodiment of the pharmaceutical composition provided herein, thepharmaceutical composition comprises about 30-70% of at least one sugarby weight of composition. In a further embodiment, the pharmaceuticalcomposition comprises about 50-60% of lactose by weight of composition.In a further embodiment, the pharmaceutical composition comprises about50-60% of lactose monohydrate by weight of composition. In a preferredembodiment, the pharmaceutical composition comprises about 55-56% oflactose monohydrate by weight of composition.

Suitable cellulose-derivative excipients include, but are not limitedto, microcrystalline cellulose, microfine cellulose, powdered cellulose,methyl cellulose, hydroxyethyl cellulose, hydroxymethyl cellulose,hydroxypropyl cellulose, and hydroxypropyl methylcellulose. In apreferred embodiment, the cellulose-based excipient is microcrystallinecellulose.

In one embodiment of the pharmaceutical composition provided herein, thepharmaceutical composition comprises about 20-40% cellulose-derivativeexcipient by weight of composition. In a further embodiment, thepharmaceutical composition comprises about 20-40% microcrystallinecellulose by weight of composition. In a further embodiment, thepharmaceutical composition comprises about 30-40% microcrystallinecellulose by weight of composition. In a preferred embodiment, thepharmaceutical composition comprises about 30-36% microcrystallinecellulose by weight of composition.

The pharmaceutical composition can, for example, be in a form suitablefor oral administration in a dosage unit form, such as a tablet,capsule, pill, powder, sustained release formulations, solution,suspension, for parenteral injection as a sterile solution, suspensionor emulsion, for topical administration as an ointment or cream or forrectal administration as a suppository. The pharmaceutical compositioncan be in unit dosage forms suitable for single administration ofprecise dosages. It will be appreciated that the unit content of acombination partner contained in an individual dose of each dosage formneed not in itself constitute an effective amount since the necessaryeffective amount may be reached by administration of a plurality ofdosage units.

Methods of preparing various pharmaceutical compositions with a specificamount of active compound are known, or will be apparent, to thoseskilled in this art. For examples, see Remington's PharmaceuticalSciences, Mack Publishing Company, Ester, Pa., 15^(th) Edition (1975).If not indicated otherwise, the formulation of the present invention isprepared in a manner known per se, for example by means of variousconventional mixing, comminution, direct compression, granulating,sugar-coating, dissolving, lyophilizing processes, or fabricationtechniques readily apparent to those skilled in the art. Solidcompositions of a similar type may also be employed in soft and hardfilled gelatin capsules. In a preferred embodiment, the pharmaceuticalcomposition of the present invention is prepared by direct compression.

As described below, the pharmaceutical composition can compriseadditional excipients or carriers, including but not limited todisintegrants, lubricants, glidants, binders, stabilizers, and fillers,diluents, colorants, flavours and preservatives. One of ordinary skillin the art may select one or more of the aforementioned carriers withrespect to the particular desired properties of the dosage form byroutine experimentation and without any undue burden. The amount of eachcarriers used may vary within ranges conventional in the art. Thefollowing references which are all hereby incorporated by referencedisclose techniques and excipients used to formulate oral dosage forms.See The Handbook of Pharmaceutical Excipients, 4^(th) edition, Rowe etal., Eds., American Pharmaceuticals Association (2003); and Remington:the Science and Practice of Pharmacy, 20^(th) edition, Gennaro, Ed.,Lippincott Williams & Wilkins (2003). These optional additionalconventional carriers may be incorporated into the oral dosage formeither by incorporating the one or more conventional carriers into theinitial mixture or added during the mixing phases.

Examples of pharmaceutically acceptable disintegrants include, but arenot limited to, starches; clays; celluloses; alginates; gums;cross-linked polymers, e.g., cross-linked polyvinyl pyrrolidone orcrospovidone, e.g., POLYPLASDONE XL from International SpecialtyProducts (Wayne, N.J.); cross-linked sodium carboxymethylcellulose orcroscarmellose sodium (e.g., AC-DI-SOL from FMC); and cross-linkedcalcium carboxymethylcellulose; soy polysaccharides; and guar gum. Thedisintegrant may be present in an amount from about 0% to about 10% byweight of the composition. In one embodiment, the disintegrant ispresent in an amount from about 0.1-5%, or about 1-3%, or about1.5-20.5% by weight of composition.

In one embodiment, the pharmaceutical composition of the presentinvention includes the disintegrant croscarmellose sodium. In a furtherembodiment, the pharmaceutical composition of the present inventionincludes about 2% croscarmellose sodium by weight of composition.

Examples of pharmaceutically acceptable lubricants and pharmaceuticallyacceptable glidants include, but are not limited to, colloidal silicondioxide/colloidal anhydrous silica (e.g., Aerosil 200®), magnesiumtrisilicate, starches, talc, tribasic calcium phosphate, magnesiumstearate, aluminum stearate, calcium stearate, magnesium carbonate,magnesium oxide, polyethylene glycol, powdered cellulose andmicrocrystalline cellulose. The lubricant may be present in an amountfrom about 0% to about 10% by weight of the composition. In oneembodiment, the lubricant may be present in an amount from about0.1-1.5%, about 0.1-1%, or about 0.5-0.9% by weight of composition. Theglidant may be present in an amount from about 0.1-10%, about 0.1-5%, orabout 0.1-1% by weight of composition.

In one embodiment, the pharmaceutical composition of the presentinvention includes the glidant colloidal silicon dioxide/colloidalanhydrous silica. In a further embodiment, the pharmaceuticalcomposition of the present invention includes about 0.25% (by weight ofcomposition) colloidal silicon dioxide/colloidal anhydrous silica.

In another embodiment, the pharmaceutical composition of the presentinvention includes the lubricant magnesium stearate. In a furtherembodiment, the pharmaceutical composition of the present inventionincludes about 0.75% magnesium stearate by weight of composition.

In another embodiment, the pharmaceutical composition of the presentinvention includes colloidal silicon dioxide/colloidal anhydrous silicaand magnesium stearate. In a further embodiment, the pharmaceuticalcomposition of the present invention includes about 0.25% colloidalsilicon dioxide/colloidal anhydrous silica by weight of composition andabout 0.75% magnesium stearate by weight of composition.

Examples of pharmaceutically acceptable binders include, but are notlimited to, starches; celluloses and derivatives thereof, for example,microcrystalline cellulose, e.g., AVICEL PH from FMC (Philadelphia,Pa.), hydroxypropyl cellulose hydroxylethyl cellulose andhydroxylpropylmethyl cellulose METHOCEL from Dow Chemical Corp.(Midland, Mich.); sucrose; dextrose; corn syrup; polysaccharides; andgelatin. The binder may be present in an amount from about 0-50%, orabout 2-20% by weight of the composition.

Examples of pharmaceutically acceptable diluents include, but are notlimited to, confectioner's sugar, compressible sugar, dextrates,dextrin, dextrose, lactose, mannitol, microcrystalline cellulose,powdered cellulose, sorbitol, sucrose and talc. The diluent, e.g., maybe present in an amount from about 0-80%, or about 0-50%, or about 1-40%or about 1-10% by weight of the composition.

In a particular embodiment, the pharmaceutical composition furthercomprises one or more of croscarmellose sodium, magnesium stearate, andsilicon dioxide.

When aqueous suspensions or elixirs are desired for oral administrationthe active compound therein may be combined with various sweetening orflavoring agents, coloring matters or dyes and, if desired, emulsifyingagents or suspending agents, together with diluents such as water,ethanol, propylene glycol, glycerin, or combinations thereof.

In a particular embodiment, the pharmaceutical composition comprisesabout 5-11% crystallized Compound A by weight of composition, about55-56% lactose monohydrate by weight of composition, and about 30-36%microcrystalline cellulose by weight of composition.

In another embodiment, the composition comprises about 5-11% ofcrystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide, about 55-56% of lactose monohydrate,about 30-36% of microcrystalline cellulose, by weight of composition,about 1.5-2.5% of croscarmellose sodium, about 0.5-0.9% of magnesiumstearate, and about 0.1-1% percent colloidal silicon dioxide/colloidalanhydrous silica, by weight of composition.

In another embodiment, the composition comprises about 5-11%crystallized Compound A by weight of composition, about 55-56% lactosemonohydrate by weight of composition, about 30-36% microcrystallinecellulose by weight of composition, about 2% croscarmellose sodium byweight of composition, about 0.75 percent magnesium stearate by weightof composition, and about 0.25 percent colloidal silicondioxide/colloidal anhydrous silica by weight of composition.

In another embodiment of the pharmaceutical composition, thepharmaceutical composition is in tablet form. In still anotherembodiment, the tablet is a coated tablet.

In one aspect, the pharmaceutical composition of the present inventionfurther comprises at least one additional therapeutic agent. Examples ofadditional therapeutic agents include, but are not limited to,chemotherapeutic agents or anti-tumor agents, such as mitoticinhibitors, alkylating agents, anti-metabolites, intercalatingantibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes,topoisomerase inhibitors, biological response modifiers, anti-hormones,angiogenesis inhibitors, and anti-androgens.

In one embodiment, the additional therapeutic agent is selected from thegroup consisting of paclitaxel or everolimus.

It is understood that the crystallized Compound A and the additionalagent may be administered in the same or different dosage unit forms.Further, such combination partners may be administered simultaneously,separately or sequentially.

Methods of Treating Proliferative Disease with Crystallized Compound A

In one aspect, the invention provides a method of treating aproliferative disease, particularly cancer, in a subject, comprisingadministering to the subject an effective amount of a crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (“Compound A”). It is understood thatcrystallized Compound A is in crystalline form produced by thecrystallization process described above.

In a further aspect, the invention provides a method of treating aproliferative disease, particularly cancer, in a subject, comprisingadministering to the subject an effective amount a pharmaceuticalcomposition comprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (“Compound A”), at least one sugar, and atleast one cellulose-derivative excipient. It is understood thatcrystallized Compound A is in crystalline form produced by thecrystallization process described above.

It is further understood that the present invention includes a method oftreating a proliferative disease, particularly cancer, in a subject byadministering to a subject an effect amount of a pharmaceuticalcomposition comprising crystallized Compound A as provided in eachembodiment set forth above, which is herein incorporated by reference.

In accordance with the method of treatment of the present invention, thecrystallized Compound A is administered to a subject in need thereof ina therapeutically effective amount.

Crystallized Compound A may be administered to a suitable subject dailyin single or divided doses at an effective dosage in the range of about0.001 to about 100 mg per kg body weight per day, preferably about 1 toabout 35 mg/kg/day, in single or divided doses. For a 70 kg human, thiswould amount to about 0.05 to 7 g/day, preferably about 0.05 to about2.5 g/day.

The crystallized Compound A and/or the pharmaceutical compositions ofthe present invention may be used for treatment of proliferativediseases which are cancerous or non-cancerous. It is expressly preferredfor treatment of cancer.

“Cancer” that can be treated using the pharmaceutical compositionprovided herein refers to cellular-proliferative disease states,including but not limited to solid or liquid tumors. Examples of cancerssuitable for treatment in accordance with the present invention include,but are not limited to, lung cancer, bone cancer, CMML, pancreaticcancer, skin cancer, cancer of the head and neck, melanoma, intrauterinecancer, ovarian cancer, colon cancer, rectal cancer, anal cancer,stomach or gastric cancer, breast cancer, testicular cancer, gynecologictumors (e.g., uterine sarcomas, carcinoma of the fallopian tubes,carcinoma of the endometrium, carcinoma of the cervix, carcinoma of thevagina or carcinoma of the vulva), Hodgkin's disease, esophageal cancer,cancer of the small intestine, cancer of the endocrine system (e.g.,thyroid cancer, parathyroid cancer, or adrenal gland cancer), softtissue sarcoma, cancer of the urethra, penile cancer, prostate cancer,chronic or acute leukemia, solid tumors of childhood, lymphocyticlymphomas, bladder cancer, cancer of the kidney or ureter (e.g., renalcell carcinoma), carcinoma of the renal pelvis, biliary cancer, braincancer, bladder cancer, squamous cell, peritoneal cancer, or neoplasmsof the central nervous system (e.g, primary CNS lymphoma, spinal axistumors, brain stem gliomas or pituitary adenomas). In one embodiment,the cancer is a solid tumor. In a particular embodiment, the solid tumoris metastatic or unresectable.

In one embodiment, the cancer is cancer such as brain, lung, squamouscell, bladder, gastric, pancreatic, breast, head, neck, renal, kidney,ovarian, prostate, colon, rectal, esophageal, testicular, gynecologicalor thyroid cancer.

In one embodiment, the cancer is lung cancer, squamous cell cancer,pancreatic cancer, breast cancer, head cancer, neck cancer, coloncancer, rectal cancer or melanoma.

In a particular embodiment, provided herein is a method of treatingmelanoma, pancreatic cancer, ovarian cancer, carcinoma of the fallopiantubes, peritoneal cancer, biliary cancer, colon cancer, or rectal cancerin a subject in need thereof, comprising administering to the subjectthe crystallized Compound A or the pharmaceutical composition providedherein.

In another embodiment, the proliferative disease is a non-cancerousproliferative disorder such as benign hyperplasia of the skin (e.g.,psoriasis), restenosis, prostate (e.g., benign prostatic hypertrophy(BPH), or Noonan Syndrome. Examples of additional therapeutic agentsinclude, but are not limited to chemotherapeutic agents or anti-tumoragents, such as mitotic inhibitors, alkylating agents, anti-metabolites,intercalating antibiotics, growth factor inhibitors, cell cycleinhibitors, enzymes, topoisomerase inhibitors, biological responsemodifiers, anti-hormones, angiogenesis inhibitors, and anti-androgens.

In one embodiment, the additional therapeutic agent is selected from thegroup consisting of paclitaxel or everolimus.

It is understood that the crystallized Compound A and the additionalagent may be administered in the same or different dosage unit forms.Further, such combination partners may be administered simultaneously,separately or sequentially.

In one aspect, the invention provides the use of crystallized Compound Afor the preparation of a medicament suitable for treatment of aproliferative disease, particularly cancer. It is understood thatcrystallized Compound A is in crystalline form produced by thecrystallization process described above. In one embodiment, theproliferative disease is a solid tumor. In a further embodiment, theproliferative disease is a cancer as set forth above.

In another aspect, the invention provides the use of the pharmaceuticalcomposition of the present invention for the preparation of a medicamentsuitable for treatment of a proliferative disease, particularly cancer.In one embodiment, the proliferative disease is a solid tumor. In afurther embodiment, the proliferative disease is a cancer as set forthabove.

In a particular embodiment of each aspect, the proliferative disease isselected from melanoma, pancreatic cancer, ovarian cancer, carcinoma ofthe fallopian tubes, peritoneal cancer, biliary cancer, colon cancer, orrectal cancer.

In one aspect, the invention provides crystallized Compound A for use inthe treatment of a proliferative disease, particularly cancer. It isunderstood that crystalline Compound A is in crystalline form producedby the crystallization process described above. In one embodiment, theproliferative disease is a solid tumor. In a further embodiment, theproliferative disease is a cancer as set forth above.

In another aspect, the invention provides the pharmaceutical compositionof the present invention, as described above and incorporated herein byreference, for use in the treatment of a proliferative disease,particularly cancer. In one embodiment, the proliferative disease is asolid tumor. In a further embodiment, the proliferative disease is acancer as set forth above.

In a particular embodiment of each aspect, the proliferative disease isselected from melanoma, pancreatic cancer, ovarian cancer, carcinoma ofthe fallopian tubes, peritoneal cancer, biliary cancer, colon cancer, orrectal cancer.

The invention is illustrated further by the following examples which arenot to be construed as limiting the invention in scope or spirit to thespecific procedures described in them.

The starting materials and various intermediates may be obtained fromcommercial sources, prepared from commercially available organiccompounds, or prepared using well known synthetic methods.

Representative examples of the methods and formulations of the inventionare set forth below. These examples are not, however, intended to limitthe scope of the invention in any way.

EXAMPLES Abbreviations

The following abbreviations are used in the text:

CDI—1,1′-carbonyldiimidazole

DMF—N,N-dimethylformamide

KOTMS—potassiumtrimethylsilanolate

THF—tetrahydrofuran

EKNS—activated charcoal

CEFOK—microcrystalline cellulose

Example 1. Preparation of6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid methyl ester

In an inertized (N₂) reaction vessel at internal temperature 20° C. andunder exclusion of humidity and air, Compound 1 (1.0 eq.) and Compound 2(1.2 eq.) are reacted in the presence of cesium carbonate (2.4 eq.),tris(dibenzylidenaceton) dipalladium(0) (0.035 eq.) and Xantphos (0.07eq.) in a mixture of toluene and 1,4-dioxane at internal temperature of99° C. After 8 hours, the mixture is cooled to internal temperature of60° C.

Subsequently, dimethylformamide (DMF), filter aid (CEFOK) and activatedcharcoal (EKNS) are added, and the mixture is stirred and cooled tointernal temperature of 35° C. The solids are filtered off and washedwith a mixture of dimethylformamide and toluene. To the filtrate, whichcontains the product Compound 3, is introduced at internal temperatureof 25° C. hydrogen chloride gas (CLC) whereupon the HCl salt of Compound3 crystallizes. The palladium residue mainly remains in solution. Afterwarming to 60° C. and cooling to 0° C., the solids are filtered using acentrifuge and are washed with a mixture of toluene anddimethylformamide.

The damp Compound 3 HCl salt is charged to a reactor (equipped with pHprobe) together with dimethylformamide and is heated to 60° C. By addinga 4 wt % of aqueous tripotassium phosphate solution, the pH is adjustedto a pH range of 6.8-7.6 (with a target of pH 7.2) while Compound 3crystallizes as free base. After cooling to 22° C. and stirring, thesolids are filtered using a centrifuge and are washed with drinkingwater. The moist solids are dried at 50° C. under vacuum to give dry,crude Compound 3.

In order to remove residual palladium, dry, crude Compound 3 isdissolved in dimethylformamide at internal temperature of 60° C. andstirred together with Smopex-234 (commercially available from JohnsonMatthey) and activated charcoal for 90 minutes. The solids are filteredoff at internal temperature of 60° C. and are washed withdimethylformamide. To the filtrate are added drinking water and Compound3 seed crystals. More drinking water is added while Compound 3crystallizes. After cooling to internal temperature of 20° C., thesolids are filtered using a centrifuge and are washed with a mixture ofdeionized water and dimethylformamide and with deionized water. Themoist solids are dried at 50° C. under vacuum, providing6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid methyl ester (Compound 3).

Example 2. Preparation of6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide

A. “One-Pot” Synthesis

In an inertized reaction vessel at internal temperature 20-25° C. undernitrogen,6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid methyl ester (Compound 3, 1.0 eq.) is added to a mixture of DMF andTHF. To this slurry, a solution of potassium trimethylsilanolate (1.05eq.) in THF is added to the mixture at internal temperature of 25° C.over a period of about 40 minutes, and the resulting mixture is stirredfor about 1 hour, providing a potassium salt solution of Intermediate 1.A THF/methanol mixture is then sequentially distilled off from themixture at 85-120° C. during about 2 hours.

The potassium salt solution is then added to a suspension of CDI (1.25eq.) and imidazole hydrochloride (1.40 eq.) in THF at internaltemperature of 25° C. over a period of about 1 hour. The resultingmixture is then stirred for approximately 1 hour at 50° C., and thefollowing imidazolide intermediate is formed:

The imidazolide intermediate is not further isolated.

Subsequently, 1.2 eq. of O-(2-tert-butoxyethyl)hydroxylamine (Compound4, CAS No. 1023742-13-3, available from suppliers such as HuhuTechnology, Inc.®) is added over a period of about 30 minutes at 50° C.and stirred for 1.5 hours. Demineralized water is then added at 50° C.,producing a precipitate. After cooling to 20° C. and stirring for about3-16 hours, the slurry is filtered off, washed with THF/demineralizedwater (1:2) in 2 portions and with demineralized water in threeportions, and dried at 50° C./<70 mbar for about 17 hours, providing6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide (Compound 5) as monohydrate.

B. A Synthesis Method with Isolation of the Intermediate of Step a) fromthe Reaction Mixture of Step a) Prior to the Reaction of Step b)

Alternatively,6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide (Compound 5) can be made by thesynthesis method as shown below. Compound 3, which is a methyl ester, isfirst converted to a carboxylic acid, which is then isolated by acrystallization to form Compound 6. Compound 6 is then coupled withCompound 4 to form Compound 5 as monohydrate. The crystallization stepin this method removes starting materials such as Compound 1, processimpurities, and the dba ligand from the prior catalyst before thecoupling reaction with Compound 4, and at the same time maintains theoverall yield of the synthesis.

6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid

In an inertized (N₂) reaction vessel at internal temperature of 60° C.,Compound 3 (1.0 eq.) is dissolved in DMF and stirred with a fiber, whichis sold under the trademark SMOPEX 234, and activated charcoal for theremoval of palladium to not more than 100 ppm. The fiber and activatedcharcoal are removed by filtration at 60° C. and washed with DMF.

The filtrate (containing Compound 3) is transferred to a secondinertized (N₂) reaction vessel and cooled to an internal temperature of30° C. A thin suspension can form at this point of time. 30% sodiumhydroxide (1.1 eq.) and water (for rinsing) are added, and the resultingreaction mixture is vigorously stirred for 3 hours at an internaltemperature of 30° C. The methyl ester is saponified. Conversion ischecked by an IPC (HPLC). As soon as the IPC criterion is met, a filteraid, which is sold under the trademark HYFLO, is added. The mixture isstirred for 15 minutes and then filtered at 30° C. via a plate filterand polish filter to a third reaction inertized (N₂) vessel.

An aqueous HCl solution 7.5% is added to the clear filtrate in the thirdvessel at an internal temperature of 30° C. until a pH value of 8 isreached. Then the solution is seeded at an internal temperature of 30°C. with Compound 6, and an aqueous HCl solution 7.5% is added undervigorous stirring until a pH value of pH 2.8 is reached. The productgradually crystallizes. The suspension is cooled over 60 min to aninternal temperature of 25° C. and water is added. The suspension isstirred for at least 4 hours at an internal temperature of 25° C.

The resulting solid is collected by centrifugation or filtration. Thefilter cake is first washed with DMF/water 1:1 (w/w) and then withwater, discharged and dried in a vacuum at 50° C. The water content iscontrolled by IPC. The crystalline product Compound 6 is discharged assoon as the IPC criterion is met.

6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide

An inertized (N₂) reaction vessel is charged with Compound 6 (1.0 eq.),DMF, and THF at room temperature. The suspension is heated to 25° C.under stirring with flow of nitrogen. After CDI (1.13 eq.) is added, thesuspension can get thinner and slight evolution of gases can beobserved. After the suspension finally becomes a solution, it is thenmonitored by IPC (HPLC).

As soon as the IPC (HPLC) criterion is met, the reaction mixture isheated to 50° C. over 20 minutes and imidazole hydrochloride (0.3 eq.)is added, forming a solution of Intermediate 2.

To the solution of Intermediate 2, Compound 4 (1.3 eq.) is added over 60minutes at internal temperature of 50° C. under stirring at a speed of300 rpm with flow of nitrogen. As soon as the IPC (HPLC) criterion ismet, the mixture is cooled to 20-25° C. over 30 minutes. The mixture isthen stored at ambient temperature overnight under nitrogen withoutstirring. DMF is added to the mixture followed by heating it to 50° C.over 30 minutes. Complete conversion of Intermediate 2 to Compound 5 isconfirmed by IPC (HPLC).

Water is added to the mixture at internal temperature of 50° C. over 20minutes. Then the solution is seeded with Compound 5. After stirring at50° C. for 60 minutes, more water is added to the suspension at 50° C.over 90 minutes. After vigorous stirring, the suspension is cooled to20° C. over 2 hours and filtered. The filter cake is washed twice withTHF/water (v/v: 1:2) at 20° C., and twice with water at 20° C. Finally,the filter cake is dried at 50° C. under vacuum to provide6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide (Compound 5) as monohydrate.

Example 3. Preparation of6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (Compound A)

6-(4-Bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid-(2-tert-butoxyethoxy)-amide (Compound 5) monohydrate is added in 3portions to a premixed solution of Acetonitrile and excess Phosphoricacid (85% aqueous solution) at internal temperature 20-25° C. Afterstirring for about 15 minutes, the suspension is heated to internaltemperature 50-53° C. The suspension is maintained at this temperaturefor 6 hours, cooled to internal temperature 20-25° C. The mixture isthen heated to internal temperature 35-37° C. and diluted withEthanol-Water (3:1 v/v). EKNS and CEFOK are added, the reaction mixtureis stirred approximately 15 minutes and filtered over a funnel coatedwith CEFOK. The filtrate is cooled to approximately 30° C. 3 N aqueouspotassium hydroxide (KOH) is added to the cooled filtrate over a periodof 90 minutes until a pH-value of about 8.1 is reached. The suspensionis heated to internal temperature 60-63° C., stirred at this temperaturefor a period of about 2 hours, cooled to 20-23° C. over a period ofabout 45 minutes, filtered over a funnel, and dried at 50° C. pressure<100 mbar over a period of about 17 hours, providing6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (Compound A) as a white powder.

Example 4. Preparation of Crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide (Compound A)

In a dry vessel at room temperature, Compound A is added to a premixedsolvent solution of methanol/THF/water (35/35/30 w/w). The suspension isheated to internal temperature 53-55° C., and the resulting solution ishot filtered by deep and membrane filtration (via a paper filter andPTFE membrane) at internal temperature 53-56° C. The clear solution isstirred and cooled to 47-48° C., and the seed crystals suspension (i.e.,seed crystals of crystallized Compound A in water, 10% m/m) is added(0.2 to 0.5% of crystallized Compound A expected yield mass). Afterabout 20 minutes, water is slowly added within 25 hours (33.3% within 15hours and 66.6% within 10 hours with at least 10 minute stirring afteraddition of water) to obtain a final ratio of methanol/THF/water(20/20/60 w/w). After the water is added, the suspension is cooled downto internal temperature 3-5° C. within 10 hours and stirred for 0.5hours. The white suspension is filtered over a sinter glass nutsche (75ml, diameter=6 cm, pore 3) suction filter and washed once with ice coldmethanol/THF/water (15/15/70 w/w at 2-4° C.), and two times with icecold water (2-4° C.). Drying takes place in a vacuum oven dryer at 20°C. for 10 hours, and then at 40° C. for 10 hours, and then at 60° C. forat least 12 hours with pressure <10 mbar, providing crystallizedCompound A.

Example 5. Pharmaceutical Composition

Crystallized Compound A is formulated as indicated in Table 1:

TABLE 1 Formulation Form 1 Form 1 Form 2 Form 2 (% by (in mg/ (% by (inmg/ weight) unit) weight) unit) Tablet core Crystallized Active 6.25% 1510.00%  15.00 Drug Substance Compound A Lactose Filler 55.63%  133.555.62%  83.43 monohydrate Microcrystalline Filler 35.13%  84.3 31.37% 47.06 cellulose Croscarmellose Disinte- 2.00% 4.8 2.00% 3 Sodium grantMagnesium Lubricant 0.75% 1.8 0.75% 1.13 Stearate Colloidal SiliconGlidant 0.25% 0.6 0.25% 0.38 Dioxide/Silica, colloidal anhydrous(e.g.,Aerosil 200 ©) TOTAL: 240 150 Tablet coating Tablet core (from  100% 100% above) Opadry II Film coat 3.50% 8.4 3.50% 8.4 (Yellow) ®**Sterile water for Solvent — — irrigation*** * The weight of the drugsubstance is taken with reference to the dried substance (100%) on thebasis of assayed value. The difference in weight is adjusted by theamount of lactose monohydrate. **The Opadry II is combined with thesterile water to make a 12% w/w Opadry II (85F) film coat suspension,which is then sprayed onto the core tablet. ***Removed during processingUpon mixing of the tablet core components, the pharmaceuticalcomposition is converted into a tablet form by direct compression. Theformed tablet may be further coated with the tablet coating providedabove.

What is claimed is:
 1. A method of treating melanoma in a patient inneed thereof, the method comprising: (a) detecting a melanoma in saidpatient that is a BRAF-mutant melanoma; and (b) administering to saidpatient a therapeutically effective amount of a pharmaceuticalcomposition comprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide.
 2. The method of claim 1, wherein saidBRAF-mutant melanoma is BRAFV600-mutant melanoma.
 3. The method of claim1, further comprising treating said patient with at least one additionaltherapeutic agent selected from the group consisting of mitoticinhibitors, alkylating agents, anti-metabolites, intercalatingantibiotics, growth factor inhibitors, cell cycle inhibitors, enzymeinhibitors, topoisomerase inhibitors, biological response modifiers,anti-hormones, angiogenesis inhibitors, and anti-androgens.
 4. Themethod of claim 3, wherein said pharmaceutical composition comprisingcrystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide and said additional therapeutic agent areadministered simultaneously.
 5. The method of claim 4, wherein saidBRAF-mutant melanoma is BRAFV600-mutant melanoma.
 6. The method of claim3, wherein said pharmaceutical composition comprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide and said additional therapeutic agent areadministered separately.
 7. The method of claim 6, wherein saidBRAF-mutant melanoma is BRAFV600-mutant melanoma.
 8. The method of claim1, wherein said pharmaceutical composition comprising crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide is formulated for oral administration. 9.The method of claim 8, wherein said pharmaceutical composition isformulated as a tablet.
 10. The method according to claim 9, whereinsaid tablet further comprises lactose monohydrate, microcrystallinecellulose, colloidal silicon dioxide, croscarmellose sodium, andmagnesium stearate.
 11. The method according to claim 9, wherein saidtablet comprises approximately 15 mg crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide.
 12. The method according to claim 10,wherein said tablet comprises approximately 15 mg crystallized6-(4-bromo-2-fluorophenylamino)-7-fluoro-3-methyl-3H-benzoimidazole-5-carboxylicacid (2-hydroxyethyoxy)-amide.